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The Supraspliceosome - A Multi-Task Machine for Regulated Pre-mRNA Processing in the Cell Nucleus.

Shefer K, Sperling J, Sperling R - Comput Struct Biotechnol J (2014)

Bottom Line: The supraspliceosome provides a platform for coordinating the numerous processing steps that the pre-mRNA undergoes: 5' and 3'-end processing activities, RNA editing, constitutive and alternative splicing, and processing of intronic microRNAs.Notably, changes in these regulatory processing activities are associated with human disease and cancer.These findings emphasize the supraspliceosome as a multi-task master regulator of pre-mRNA processing in the cell nucleus.

View Article: PubMed Central - PubMed

Affiliation: Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel.

ABSTRACT
Pre-mRNA splicing of Pol II transcripts is executed in the mammalian cell nucleus within a huge (21 MDa) and highly dynamic RNP machine - the supraspliceosome. It is composed of four splicing active native spliceosomes, each resembling an in vitro assembled spliceosome, which are connected by the pre-mRNA. Supraspliceosomes harbor protein splicing factors and all the five-spliceosomal U snRNPs. Recent analysis of specific supraspliceosomes at defined splicing stages revealed that they harbor all five spliceosomal U snRNAs at all splicing stages. Supraspliceosomes harbor additional pre-mRNA processing components, such as the 5'-end and 3'-end processing components, and the RNA editing enzymes ADAR1 and ADAR2. The structure of the native spliceosome, at a resolution of 20 Å, was determined by cryo-EM. A unique spatial arrangement of the spliceosomal U snRNPs within the native spliceosome emerged from in-silico studies, localizing the five U snRNPs mostly within its large subunit, and sheltering the active core components deep within the spliceosomal cavity. The supraspliceosome provides a platform for coordinating the numerous processing steps that the pre-mRNA undergoes: 5' and 3'-end processing activities, RNA editing, constitutive and alternative splicing, and processing of intronic microRNAs. It also harbors a quality control mechanism termed suppression of splicing (SOS) that, under normal growth conditions, suppresses splicing at abundant intronic latent 5' splice sites in a reading frame-dependent fashion. Notably, changes in these regulatory processing activities are associated with human disease and cancer. These findings emphasize the supraspliceosome as a multi-task master regulator of pre-mRNA processing in the cell nucleus.

No MeSH data available.


Related in: MedlinePlus

The supraspliceosome model. (A, B) Schematic models of the supraspliceosome in which the pre-mRNA (introns in blue, exons in red) is connecting four native spliceosomes. The supraspliceosome presents a platform onto which the exons can be aligned and splice junctions can be checked before splicing occurs. (A) The pre-mRNA that is not being processed is folded and protected within the cavities of the native spliceosome. (B, C) When a staining protocol that allows visualization of nucleic acids was used, RNA strands and loops bound with proteins were seen emanating from the supraspliceosomes [16]. (B) Under these conditions the RNA kept in the cavity is proposed to unfold and loop-out. In the looped-out scheme an alternative exon is depicted in the upper left corner. Adapted from Azubel et al. [19]. (C) STEM dark field images of supraspliceosomes [16] stained with a protocol that allows visualization of nucleic acids. Adapted from Muller et al. [16]. Bar represents 10 nm.
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f0015: The supraspliceosome model. (A, B) Schematic models of the supraspliceosome in which the pre-mRNA (introns in blue, exons in red) is connecting four native spliceosomes. The supraspliceosome presents a platform onto which the exons can be aligned and splice junctions can be checked before splicing occurs. (A) The pre-mRNA that is not being processed is folded and protected within the cavities of the native spliceosome. (B, C) When a staining protocol that allows visualization of nucleic acids was used, RNA strands and loops bound with proteins were seen emanating from the supraspliceosomes [16]. (B) Under these conditions the RNA kept in the cavity is proposed to unfold and loop-out. In the looped-out scheme an alternative exon is depicted in the upper left corner. Adapted from Azubel et al. [19]. (C) STEM dark field images of supraspliceosomes [16] stained with a protocol that allows visualization of nucleic acids. Adapted from Muller et al. [16]. Bar represents 10 nm.

Mentions: The relatively uniform mass of the supraspliceosome (21.1 ± 1.6 MDa; n = 400) [16], likely reflects the presence of a general basic structure. Using a positive staining protocol, which allowed visualization of nucleic acids, we could show strands and loops of RNA emanating from positively stained supraspliceosomes [16] (see Fig. 3C), indicating that the RNA is loosely bound and therefore accessible for probing. Furthermore, fibers, presumably the pre-mRNA covered with proteins, through which the native spliceosomes are interconnected were observed by 2-D image restoration of ice-embedded particles [28]. These observations support our working hypothesis that the native spliceosomes are connected by the pre-mRNA. Within the supraspliceosome, the native spliceosomes are arranged such that their small subunits reside in its center. This configuration allows communication between the native spliceosomes [30].


The Supraspliceosome - A Multi-Task Machine for Regulated Pre-mRNA Processing in the Cell Nucleus.

Shefer K, Sperling J, Sperling R - Comput Struct Biotechnol J (2014)

The supraspliceosome model. (A, B) Schematic models of the supraspliceosome in which the pre-mRNA (introns in blue, exons in red) is connecting four native spliceosomes. The supraspliceosome presents a platform onto which the exons can be aligned and splice junctions can be checked before splicing occurs. (A) The pre-mRNA that is not being processed is folded and protected within the cavities of the native spliceosome. (B, C) When a staining protocol that allows visualization of nucleic acids was used, RNA strands and loops bound with proteins were seen emanating from the supraspliceosomes [16]. (B) Under these conditions the RNA kept in the cavity is proposed to unfold and loop-out. In the looped-out scheme an alternative exon is depicted in the upper left corner. Adapted from Azubel et al. [19]. (C) STEM dark field images of supraspliceosomes [16] stained with a protocol that allows visualization of nucleic acids. Adapted from Muller et al. [16]. Bar represents 10 nm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4232567&req=5

f0015: The supraspliceosome model. (A, B) Schematic models of the supraspliceosome in which the pre-mRNA (introns in blue, exons in red) is connecting four native spliceosomes. The supraspliceosome presents a platform onto which the exons can be aligned and splice junctions can be checked before splicing occurs. (A) The pre-mRNA that is not being processed is folded and protected within the cavities of the native spliceosome. (B, C) When a staining protocol that allows visualization of nucleic acids was used, RNA strands and loops bound with proteins were seen emanating from the supraspliceosomes [16]. (B) Under these conditions the RNA kept in the cavity is proposed to unfold and loop-out. In the looped-out scheme an alternative exon is depicted in the upper left corner. Adapted from Azubel et al. [19]. (C) STEM dark field images of supraspliceosomes [16] stained with a protocol that allows visualization of nucleic acids. Adapted from Muller et al. [16]. Bar represents 10 nm.
Mentions: The relatively uniform mass of the supraspliceosome (21.1 ± 1.6 MDa; n = 400) [16], likely reflects the presence of a general basic structure. Using a positive staining protocol, which allowed visualization of nucleic acids, we could show strands and loops of RNA emanating from positively stained supraspliceosomes [16] (see Fig. 3C), indicating that the RNA is loosely bound and therefore accessible for probing. Furthermore, fibers, presumably the pre-mRNA covered with proteins, through which the native spliceosomes are interconnected were observed by 2-D image restoration of ice-embedded particles [28]. These observations support our working hypothesis that the native spliceosomes are connected by the pre-mRNA. Within the supraspliceosome, the native spliceosomes are arranged such that their small subunits reside in its center. This configuration allows communication between the native spliceosomes [30].

Bottom Line: The supraspliceosome provides a platform for coordinating the numerous processing steps that the pre-mRNA undergoes: 5' and 3'-end processing activities, RNA editing, constitutive and alternative splicing, and processing of intronic microRNAs.Notably, changes in these regulatory processing activities are associated with human disease and cancer.These findings emphasize the supraspliceosome as a multi-task master regulator of pre-mRNA processing in the cell nucleus.

View Article: PubMed Central - PubMed

Affiliation: Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel.

ABSTRACT
Pre-mRNA splicing of Pol II transcripts is executed in the mammalian cell nucleus within a huge (21 MDa) and highly dynamic RNP machine - the supraspliceosome. It is composed of four splicing active native spliceosomes, each resembling an in vitro assembled spliceosome, which are connected by the pre-mRNA. Supraspliceosomes harbor protein splicing factors and all the five-spliceosomal U snRNPs. Recent analysis of specific supraspliceosomes at defined splicing stages revealed that they harbor all five spliceosomal U snRNAs at all splicing stages. Supraspliceosomes harbor additional pre-mRNA processing components, such as the 5'-end and 3'-end processing components, and the RNA editing enzymes ADAR1 and ADAR2. The structure of the native spliceosome, at a resolution of 20 Å, was determined by cryo-EM. A unique spatial arrangement of the spliceosomal U snRNPs within the native spliceosome emerged from in-silico studies, localizing the five U snRNPs mostly within its large subunit, and sheltering the active core components deep within the spliceosomal cavity. The supraspliceosome provides a platform for coordinating the numerous processing steps that the pre-mRNA undergoes: 5' and 3'-end processing activities, RNA editing, constitutive and alternative splicing, and processing of intronic microRNAs. It also harbors a quality control mechanism termed suppression of splicing (SOS) that, under normal growth conditions, suppresses splicing at abundant intronic latent 5' splice sites in a reading frame-dependent fashion. Notably, changes in these regulatory processing activities are associated with human disease and cancer. These findings emphasize the supraspliceosome as a multi-task master regulator of pre-mRNA processing in the cell nucleus.

No MeSH data available.


Related in: MedlinePlus